Methamphetamine induces dopaminergic neurite degeneration, oxidative damage, and a-synuclein (aSyn) aggregation in the nigrostriatal system. Similar neuropathological features are evident in the brains of individuals with Parkinson's disease (PD). Familial and sporadic PD involve dysfunction of DJ-1, a neuroprotective protein that suppresses oxidative stress and aSyn aggregation by activating various pro- survival mechanisms. Collectively, these observations suggest that DJ-1 may protect against methamphetamine neurotoxicity. The long-term objective of the proposed research is to identify cellular responses that abrogate dopaminergic neurite degeneration elicited by methamphetamine, as a way to stimulate the discovery of new therapies. The studies described in this application are focused on the role of DJ-1 in alleviating methamphetamine neurotoxicity. This question will be addressed with the following specific aims: (i) determine whether DJ-1 inhibits methamphetamine-induced neurite loss;(ii) assess whether DJ-1 suppresses oxidative stress and 1-synuclein aggregation in methamphetamine-treated cells;and (iii) determine whether DJ-1-mediated protection against methamphetamine neurotoxicity requires expression of the protein in glia and/or neurons. Primary midbrain cultures expressing human wild-type DJ-1 from an adenoviral construct will be exposed to methamphetamine, and the percentage of dopaminergic neurons without processes will be determined immunocytochemically. The cells will also be characterized in terms of a range of DJ-1 protective responses. The effect of subcellular localization will be investigated by measuring the neuroprotective activity of engineered DJ-1 variants preferentially targeted to the cytosol, mitochondria, or nucleus. The role of cell-type-specific DJ-1 expression will be examined by monitoring protective pathways in cultures that express DJ-1 selectively in neurons, astrocytes, or microglia. This project will advance our understanding of cellular responses that alleviate methamphetamine neurotoxicity. Ultimately, the insights from this study will stimulate the development of strategies to treat individuals exposed to methamphetamine via activation of neuroprotective pathways modulated by DJ-1. PUBLIC HEALTH RELEVANCE: This project is designed to assess neuroprotective mechanisms by which the protein DJ-1 suppresses the loss of neuronal processes triggered by methamphetamine. The insights from these studies will stimulate the development of therapeutic strategies to alleviate methamphetamine neurotoxicity via upregulation of DJ-1 protective responses.